Research directed toward the long term goal of designing in vitro systems for the cultivation of Onchocerca lienalis and O. volvulus through the invertebrate and early mammalian phases of their life cycles is proposed. This research is prompted by the need for a system to produce stage specific antigens free of complicating factors from the host for the early diagnosis and prevention of onchocercid infections. The need for in vitro technology is particularly acute in the case of O. volvulus because at present, no satisfactory animal host is available for this parasite. The project will be conducted by a medical entomologist in consultation with a parasitologist and a specialist in vertebrate and invertebrate cell culture. The research plan draws heavily upon newly developed techniques for rearing black flies and artifically infecting them with Onchocerca spp. The derivation of black fly cell cultures for co-cultivation with onchocercid larvae is the first specific aim of the project. Conditions for the cultivation of invertebrate phase larvae in cell free media, isolated black fly thoracic muscles and insect cell cultures will be optimized. Control for microfilarial viability and protocols involving transplantation of larvae from vector to culture at various times during development will be stressed. Co-cultivation of onchocercid larvae with insect cells on inert three dimensional matrices will be tested for positive effect. Finally, techniques for culture of third-stage larvae of Onchocerca spp. will be perfected. In general, culture techniques will be optimized using the bovine parasite Onchocerca lienalis prior to testing with O. volvulus. Laboratory reared females of the nearctic black fly sepeies Simulium decorum, S. vittatum and S. pictipes will act as donors of onchocercid larvae and cell culture explants.